Various forms of silica and silica-based chromatographic materials are of primary importance in separating fine organic and biochemical products from one another, from by-products and from excess reagents in reaction mixtures. For instance, silica of various particle sizes (e.g., 1 to 700 .mu.m) and pore diameters (e.g., 1 nm to 1 .mu.m) is used in a very large fraction of the high-pressure liquid chromatographic separation techniques ("HPLC") in use. For certain applications (e.g., affinity chromatography) active groups which exhibit different degrees of interaction with different components of a mixture to be separated are coated on or bonded to the silica surface, usually by means of using organo-silane reagents. A particular class of modified silica-based chromatographic supports are controlled pore glass supports, which are coated with polar groups such as quarternary amine or sulfonic glycophases for use in ion-exchange chromatography. Controlled pore glass supports are also widely used in exclusion and affinity chromatography. On the other hand, despite the availability of a large variety of silica supports coated with various functional groups and molecules, materials consisting of largely pure silica are used in a large number of separations. For instance, silica gel is the most widely used support in high pressure liquid chromatography. Molecules separated on pure and coated silica surfaces include amines, amino acids, purines, nucleosides, nucleotides, aminoglycosides, peptides, proteins, nucleic acids, enzymes and other organic and biochemical molecules as well as inorganic species such as metal ions and anions.
In addition to their role in separations, silica-based supports are also used in carrying out chemical reactions using reagents which are immobilized on the grain and pore surfaces. Such reactive supports include, for instance, reducing supports to which reducing groups such as borohydride have been attached, supports which contain chelating groups to remove metal ions from solutions, and supports which contain immobilized enzymes and other biochemically active species.
In spite of their wide usefulness, silica-based supports are subject to a number of limitations. These are evident in some cases more than in others. For instance, uncoated silica supports are more susceptible to deterioration at high pH than supports coated with neutral molecules, and coarse-grained, large-pore glass supports are more durable than fine-grained, small pore gels. However, the problems encountered in the case of silica gels used for high-pressure liquid chromatography, for instance, also appear in the cases of other silica-based supports, although their extent may be smaller or they may appear after a longer period of service.
One serious problem which limits the application of silica supports is the poor durability of silica at the high pH range. Generally, mixtures of organic or biological molecules are separated by sorption on a support followed by fractional elution using eluants with varying combinations of pH and ionic strength. High pH separations are very useful in the case of proteins and other substrates. However, at pH values above 7.5 rapid silica dissolution takes place, leading to loss of activity of surface sites and to clogging due to reprecipitation at the lower parts of the column. Support deterioration becomes much more serious at elevated temperatures.
Jackson and Fisher (Ind. Res. Dev., Feb. 1983, pp. 130-33) state: "For a silica-based bonded phase, the packings are stable in the pH range of 2.0 to 7.5 Exposure to a mobile phase with a pH higher than 7.5 (less acid) causes dissolution of the silica and leads to voids in the column. The resulting chromatogram will exhibit peak broadening, and thus a loss of resolution, sensitivity (as the peak broadens, the peak height decreases, decreasing the sensitivity), and quantitative accuracy. We need to note also that a sample of high pH can cause void formation at the column head as it is introduced onto the column. This effect can be observed regardless of the pH of the column. Of course, an occasional run outside the pH range, 2.0 to 7.5, can be acceptable. But, frequent operation outside the range results in both poor chromatography and greatly reduced column life". In addition to poor column performance and short lifetime, the enhanced silica dissolution at pH values above 7.5 also causes considerable contamination of the separated products with silica. One way to overcome the problems associated with the use of silica columns at high pH environments, mentioned by Jackson and Fisher, is to replace such columns by resin-based, usually polystyrene/divinylbenzene, stationary phases. However, these materials have poor mechanical properties and are crushed at the high pressures required in typical liquid chromatography operations. Jackson and Fisher also emphasize that "even small changes in pH (e.g., by 0.5 of a pH unit) can have large effects on the shapes of chromatograms". Accordingly, even a small increase in the service range of silica-based stationary phases can result in considerable improvement of existing separation techniques and in making it possible to perform novel separations.
Another problem associated with the use of silicabased supports is the fact that the silanol surface groups often form excessively strong bonding with the organic or biochemical species being purified and this requires the use of drastic conditions (in terms of solvent polarity, pH, ionic strength and temperature) to desorb the molecule of interest. In the case of many sensitive organic structures this leads to decomposition or loss of biological activity (for instance, denaturing of proteins).
A third problem associated with the use of silica supports is due to the existence of various configurations of silicon-oxygen on silica surfaces, including isolated hydroxyl groups, pairs of hydroxyl groups attached to the same Si atoms, pairs of hydroxyl groups attached to adjacent Si atoms, and siloxane (Si-O-Si) terminal groups. Since each of these configurations has a different affinity towards organic substrates, chromatographic peaks are often quite broad and the resolution of separating similar species is poor.
It is well-known that hydrous oxides of various metal ions such as Al and Fe reduce the solubility of silica in water (U.S. Pat. No. 2,267,831) and in aqueous solutions (U.S. Pat. No. 4,332,031). This is due to the formation of a combined species (aluminosilicate, iron oxide-silicate, etc.) which has a very low solubility in water. The interaction involved in the formation of such species may consist of a chemical reaction, sorption, ion-exchange, colloid-colloid neutralization, etc. Furthermore, the rise in solubility with increasing temperature is also smaller in the case of aluminosilicates than in the case of silica. However, according to studies of the chemical durability of silicate glasses in high pH media (Ohta and Suzuki, Am. Ceram. Soc. Bull., 57, 602-604 (1978)), the addition of alumina or iron oxide to the glass results in increased corrosion rates.
U.S. Pat. No. 3,843,341 relates to thermally, stable, mechanically strong microporous glass articles with large pore volumes, surface areas, and varying pore sizes, and methods for making such articles. In particle form, such as beads, the microporous glass articles are useful as catalyst supports in applications such as petroleum catalytic refiners and motor vehicle catalytic mufflers. The mechanical strength and the dimensional stability of the microporous glass articles at elevated temperatures can be improved if the articles are preshrunk, such as by brief exposure to high temperatures, before their intended use, and can be improved even further if treated with certain metal oxides. In particular, this improvement is achieved by treating the preshrunk porous glass article with a tin solution to deposit tin oxide thereon. The deposition is effected through soaking in a tin chloride solution, drying and calcination at 700.degree. C. A similar procedure is used with other metals such as aluminum and zirconium. The deposition of the metal salt occurs during the initial drying stage performed at 100.degree. C. This causes the deposition to take place through thermal precipitation as water is evaporated rather than through sorption on the glass surface. Furthermore, the metal salt solutions used, such as solutions of tin chloride, aluminum nitrate, and zirconium oxychloride, are highly acidic and accordingly they are not suitable for sorption of the metal ion upon contact with the porous glass surface. However, tin oxide (SnO.sub.2) is a particularly desirable metal oxide to deposit on the porous glass articles of the invention. Tin oxide has been found to provide additional thermal stability to the porous glass bead. According to the patent, this is somewhat surprising in that other metal oxides such as Al.sub.2 O.sub.3, Cr.sub.2 O.sub.3 and ZrO.sub.2 do not provide analogous results and do not show improved stability over the undoped glass.
The amount of tin oxide deposited within the pores according to this patent should be about 0.5 to 10 percent, preferably about 1 to 5 percent by weight. The metal oxide doping levels shown in this patent are 1.5% SnO.sub.2, 1% Al.sub.2 O.sub.3, 1.5% Cr.sub.2 O.sub.3, 3.0% ZrO.sub.2 and 3% CuO-Cr.sub.2 O.sub.3, respectively. In all cases, the metal oxide deposition treatment follows a high-temperature preshrinking step to produce a porous glass article with improved stability in high-temperature applications.
U.S. Pat. No. 3,923,688 is a continuation-in-part of the previous patent. It details the preparation of the microporous glass articles having a catalytic coating deposited thereon, where the catalyst is a metal oxide selected from the class consisting of TiO.sub.2, V.sub.2 O.sub.5, Cr.sub.2 O.sub.3, FeO, CoO, NiO.sub.2, CuO, Al.sub.2 O.sub.3, ZrO.sub.2 and SnO.sub.2. It also mentions the possible uses of the thermally stable porous glass of the invention as an ion exchange and desalination medium, as a chromatographic support, a filter for gases and vapors and as a membrane providing an effective diffusion barrier for use in ultrafiltration and reverse osmosis. In these applications, the patent notes that porous glass offers expanded capabilities for use in systems employing high temperature and low pH which are generally not compatible with organic polymer membranes.
U.S. Pat. No. 4,178,270 describes an inorganic ion-exchanger supported on a carrier which is prepared by supporting an active component of inorganic ion exchanger, such as hydrous oxide of metal, for example, titanium, zirconium, etc., on a porous carrier such as alumina, silica, or activated carbon, where the pH of a solution in contact with the active component and the carrier is adjusted so that the active component and the carrier can have zeta potentials of opposite polarities to each other, and an inorganic ion-exchanger having a larger amount of the supported active component and firmly supporting less bleedable active component can be obtained by setting out a supporting condition on the basis of the polarity of zeta potential. This patent describes the preparation of supported hydrous titanium and zirconium oxides, but the possible use of the hydrous oxide of titanium in admixture with manganese, zinc, tin, zirconium, silicon or rare earth elements is also mentioned. According to the invention described in the patent, the carrier supports about 10% by weight of the hydrous metal oxide in the hydrolyzed state, on the basis of the carrier, and at least 5% by weight thereof, even after shaking. This invention does not describe a chromatographic material. It specifically describes an ion-exchanger suitable for recovery of useful resources in sea water, removal of impurities from high temperature boiler water in nuclear reactors or industrial boilers, removal of impurities in industrial waste water, etc. In this invention alumina is considered as a carrier rather than as a potential dopant.
U.S. Pat. No. 4,333,847 relates to the immobilization of toxic, e.g., radioactive materials, internally in a silicate glass or silica gel matrix for extremely long periods of time. Toxic materials, such as radioactive wastes containing radioactive anions, and in some cases cations, which may be in the form of liquids, or solids dissolved or dispersed in liquids or gases, are internally incorporated into a glass matrix, having hydrous organofunctionalsiloxy groups, e.g., hydrous aminoalkylsiloxy or carboxyorganosiloxy, bonded to silicon atoms of said glass and/or hydrous polyvalent metals bonded to silicon atoms of said glass through divalent oxygen linkages or otherwise immobilized therein, by a process which involves the ion exchange of said toxic, radioactive anions with hydroxyl groups attached to said organofunctionalsiloxy groups or with hydroxyl groups attached to the hydrous polyvalent metal. In the case where hydrous polyvalent metals are used, said non-radioactive cationic polyvalent metals are selected from the group consisting of -Zr.sup.3+, -Pb.sup.+, -Th.sup.3+ and -Ti.sup.3+. The processes described in the patent for attaching said metals to the glass consist of cation exchange with protons or molecular stuffing, which lead to loading the glass with high concentrations of hydrous oxides of said metals.
U.S. Pat. No. 3,677,938 relates to a chromatographic separation process using columns filled with porous silica. The porous silica used in the columns is prepared by calcination of silica gel at temperatures within the range of 400.degree. to 1000.degree. C. The silica gel used in the calcination is doped with foreign atoms including alkali metal cations (lithium, sodium, potassium and cesium) and acid anions (sulfate, phosphate, bromide, chloride and iodide). This formulation makes it possible to obtain substantially complete dehydration of the silica upon calcination, leading to enhanced mechanical strength and stability of the silica gel particles.
U.S. Pat. No. 3,722,181 relates to a process for making a chromatographic packing having a polymeric stationary phase comprising repeating units of silicon. The organic stationary phase is bonded to a substrate which contains a metal or a metal oxide where the metal has a valence of 3-5, including non-alkaline metal oxides, alumina, thoria, titania, zirconia and non-alkaline metals with an oxide skin. However, the preferred substrates are those which contain silica, such as diatomaceous earth, silica gel, glasses, sand, aluminosilicates, quartz, porous silica beads and clays. The purpose of the introduction of Si or a polyvalent metal into the substrate surface is to provide for the formation of linkage with the polymolecular organosilicon stationary phase.
U.S. Pat. No. 4,299,732 relates to amorphous aluminosilicates which are useful as catalysts. It describes the preparation of synthetic amorphous aluminosilicates by mixing under reaction conditions a source of silica such as an aqueous colloidal dispersion of silica particles, a source of alumina such as sodium aluminate prepared by dissolving alumina particles in excess sodium hydroxide solution, a source of alkali metal such as sodium hydroxide, water and one or more polyamines other than a diamine. The reaction conditions are a temperature in the range 80.degree. to 210.degree. C., a pressure in the range of 70 to 400 psig and a reaction time in the range 20 to 100 hours. The exact formulation of the aluminosilicate and the specific method of preparation have a large effect on the catalytic properties of the surface of aluminosilicate solids.
U.S. Pat. No. 4,322,310 relates to a composition comprising a chiral organic amine covalently linked via a carbamate, mercaptocarbamate, or urea linkage to a chain of atoms which in turn are covalently bound to a core support. The support described in the patent is used as a solid phase chromatographic medium in the separation of racemic mixtures. This patent illustrates, for example, the binding of a silyl group to an alumina support and having an alpha-methylbenzyl amine molecule bound through a carbamate linkage to said 3-propyl-silyl group. In general, supports mentioned in this patent include silica, alumina, glass and ceramic materials, and silylating agents are used to form a covalent bonding to the chiral composition.
U.S. Pat. No. 4,340,496 relates to an anion exchange composition that is useful in chromatographic separations comprising an inert porous particle having a tetra-substituted silane material fixedly attached by covalent bonding to the surface thereof. This patent demonstrates the use of the resulting weak anion exchange material in separating polar polyfunctional compounds such as proteins. The inert porous particle to which the silane material is attached is microparticulate silica in all the examples cited in the patent. The possible use of alumina, cross-linked dextran and cross-linked polystrene-divinylbenzene as inert porous particles is also mentioned.
U.S. Pat. No. 4,359,389 relates to the purification of human fibroblast interferon using a two-stage purification method comprising (a) subjecting an aqueous interferon solution to chromatography on porous glass beads and (b) subjecting the resulting aqueous interferon solution to chromatography on immobilized zinc chelate. The porous glass beads used in the first stage are of controlled (i.e., rather uniform) pore size between 170 and 1700 .ANG. (usually between 350 and 900 .ANG.) and a diameter which may be less uniform and may range in general between 50 and 500 .mu.m. The beads are used to absorb the interferon from an aqueous solution having a neutral or slightly alkaline pH (around 7.4), and the interferon is eluted using an elution agent at acidic pH (6 to 4) and subjected to the second stage of purification by means of immobilized zinc chelate.
Accordingly, it is an object of the present invention to provide an improved silica-based support.
A further object of the present invention is to provide a silica-based chromatographic or reactive support which is relatively free from deterioration at high pH or high temperature.
Yet a further object of the present invention is to provide a silica-based support which has good durability of silica at high pH or high temperature.
Another object of the present invention is to provide a silica-based chromatographic support for bonding organic or biochemical species in which the molecules can be desorbed without substantial decomposition or loss of biological activity.
Another object of the present invention is to provide a silica-based chromatographic or reactive support which minimizes product contamination with silicate during separation of mixtures or chemical reaction.
Yet another object of the present invention is to provide a silica-based chromatographic support which provides good resolution of separating similar species.
Another object of the present invention is to provide a method whereby a chromatographic support can be improved, for instance, to have a higher stability at high pH or high temperatures, without adversely affecting the flow characteristics or the chromatographic resolution of the support material.
Another object of the present invention is to provide a durable reactive silica-based support which can be used at high pH or at high temperature or both.
Another object of the present invention is to provide a method for making a corrosion-resistant porous siliceous article which can be used as a support for a dye, an enzyme or a dye-enzyme combination in a sensor device for measuring chemical or physical properties in aqueous media over significant time periods.